RU2536940C2 - New treatment of acute lymphoblastic leukaemia - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: present invention refers to immunology. What is presented is using Blinatumomab (MT103) for preparing a pharmaceutical composition for treating, relieving or eliminating acute lymphoblastic leukaemia (ALL), wherein Blinatumomab (MT103) transforms a MRD (minimal residual disease) positive acute lymphoblastic leukaemia ALL into the MRD-negative condition of ALL.

EFFECT: using the invention provides transforming the MRD-positive acute lymphoblastic leukaemia ALL into the MRD-negative condition of ALL molecularly that can find application in medicine in the therapy of recurrent ALL.

21 cl, 8 dwg, 1 tbl, 1 ex

Description

The present invention relates to a method for treating, reducing the intensity or elimination of acute lymphoblastic leukemia (ALL), comprising administering a pharmaceutical composition comprising the construction of a CD19 × CD3 bispecific single chain antibody to an adult patient in need thereof.

Leukemia is a clonal neoplastic proliferation of immature hematopoietic cells that are characterized by aberrant differentiation or delayed differentiation. Leukemia cells accumulate in the bone marrow, eventually replacing most of the normal hematopoietic cells. This leads to bone marrow failure and its consequences in the form of anemia, bleeding and infection. Leukemia cells circulate in the blood and other tissues throughout the body (DeVita, Hellmann, Rosenberg. Cancer: principles and practice of oncology. Eight edition. Library of Congress Cataloging-in-Publication Data, ISBN 0-781-72387-6). Acute leukemia, which can be roughly classified as either lymphoblastic or myeloblastic, can be identified phenotypically and genetically and are characterized by a rapid clinical course requiring immediate treatment. Acute leukemia arises from early hematopoietic progenitor cells. In contrast, cells of chronic leukemia have a phenotype and a characteristic biological characteristic of more mature cells (DeVita et al., In the cited place). Acute lymphoblastic leukemia (ALL) is different from lymphomas, since lymphomas are similar to more mature lymphoid cells and are usually located in the lymph nodes, spleen, or other extramedullary places before spreading to the bone marrow. Certain lymphomas, such as lymphoblastic lymphomas or Burkitt’s lymphomas, retain signs of both leukemia and lymphomas, but arise from immature progenitor cells, and require therapy similar to that used for acute lymphoblastic leukemia (ALL). However, other lymphomas can significantly spread into the blood and bone marrow and at this phase can be characterized as leukemic lymphomas, rather than true leukemia (De Vita et al., In the cited place). Acute lymphoblastic leukemia is a relatively rare malignant tumor. The total incidence of acute lymphoblastic leukemia (ALL) is 1.1 / 100000 per year. The peak frequency of occurrence occurs in childhood, constantly decreasing with age. From the age of 35 years, the incidence rate again increases, and a second peak is observed starting at the age of 80 years (2.3 / 100000 per year) (Hoelzer and Gokbuget; Der Onkologe 12 (2006); 983-1002). Although the etiology of acute lymphoblastic leukemia (ALL) is unclear, it is one of the most comprehensively studied and most characterized tumors. Subgroups of acute lymphoblastic leukemia (ALL) are determined primarily through immunophenotyping, cytogenetics, and molecular genetics. Acute lymphoblastic leukemia (ALL) from B-cell cells, accounting for 74% of cases, includes most ALL. Seventy percent of all ALL are ALL from precursor B cells, and 4% ALL are ALL from mature B cells. ALL from T-line cells refers to 26% of all ALL (Hoelzer and Gokbuget; Der Onkologe 12 (2006); 983-1002).

In early 1980, acute lymphoblastic leukemia (ALL) in adults was a rarely treatable disease, with an overall survival rate of less than 10%. After the use of adapted schemes assigned to pediatric groups, a favorable outcome increased to 30-40%. The period of stop with subsequent improvement was noted only in individual subgroups. However, over the past five years, progress has been made in the molecular diagnosis of acute lymphoblastic leukemia (ALL). Stem cell transplantation increased the favorable outcome of acute lymphoblastic leukemia (ALL) and made treatment more possible. Although various new specialized drugs are being evaluated, effective specialized therapies for acute lymphoblastic leukemia (ALL) are not yet available. The rapid diagnosis and classification of acute lymphoblastic leukemia (ALL) is becoming increasingly important for identifying prognostic and molecular genetic subgroups that will be the focus of specialized treatment (Hoelzer and Gökbuget; Hematology (2006); 133-141). The Philadelphia (Ph) chromosome, the result of reciprocal translocation leading to the fusion of the abl proto-oncogen from chromosome 9 with sequences of localization regions of the rearrangement initiation site on chromosome 22, was the first tumor-specific translocation identified. Translocation t (9; 22) is the most common genetic aberration in acute lymphoblastic leukemia (ALL) in adults. It is found in 20-30% of patients. The incidence rate increases with age, reaching 50% in patients older than 50 years. In previous clinical trials, elderly patients were underrepresented due to the identified treatment failure, but this picture is changing with the availability of promising new treatment options. It is noteworthy that it is detected almost exclusively in acute lymphoblastic leukemia from CD10 + B-progenitor cells (common B-cell ALL (c-ALL) (ordinary pre-pre-B-cell ALL) and pre-B-cell ALL); Chronic myeloid leukemia (CML) in lymphoid blast crisis may represent rare reports of its presence in ALL from T-line cells, rather than true Ph + ALL. Clinically, patients who have a variable white blood cell count, surface expression of CD19, CD10, and CD34 antigens and frequent co-expression of bone marrow related markers, such as CD13 and CD33, are at increased risk of developing meningeal leukemia. The prognosis for adult patients with Ph + ALL treated only with chemotherapy is unfavorable, with the likelihood of prolonged survival of less than 10%. Due to the sad outcome of using chemotherapy, it is now believed that allogeneic hematopoietic stem cell transplantation (HSCT) is the preferred treatment for Ph + ALL in adults. Degrees of long-term survival ranging from 12% to 65% have been reported for patients undergoing SCT in first complete remission (CR), meaning that this procedure is potentially healing. However, approximately 30% of these patients experience relapse (Ottmann and Wassmann; Hematology (2005), 118-122). The presence of leukemic cells at a level that is below the cytological limit of detection (5% of leukemic cells) is defined as “minimum residual disease” (MRD). If MRD is not detected (<10 ^-4 , i.e., less than 1 leukemia cell per 10 ⁴ bone marrow cells), complete remission at the molecular level is achieved. In recent years, using a series of studies on the characteristics of a certain group of people associated with the development of a disease in the past, it has been established that MRD in acute lymphoblastic leukemia (ALL) in adults is an independent prognostic factor already established for childhood leukemia . Diagnostic tools for MRD are polymerase chain reaction (PCR) and / or flow cytometry. Using PCR analysis, fusion transcripts, such as bcr / abl, and individual clonal rearrangements of immunoglobulin (IgH) or T cell receptor (TCR) genes can be detected. Approximately 25% of patients with minimal residual disease (MRD) determined by rearrangement include a group at a high - 94% risk of relapse within 3 years. As a rule, attenuation of MRD occurs more slowly in adults than in children. Therefore, the decision made regarding the intensification of treatment using allogeneic peripheral blood stem cell transplantation (PBSCT) is very fast after treatment induction. However, after the onset of consolidation, minimal residual disease (MRD) at any time is associated with a high risk of relapse (Bruggemann et al., Blood 107 (2006), 1116-1123; Raff et al., Blood 109 (2007), 910-915).

The treatment of adult patients with acute lymphoblastic leukemia (ALL) is becoming more or more complicated, as different treatment protocols are introduced for different subtypes of the disease, reflecting the effort to optimally select therapy for the categories of diseases according to the specific risk. Recent improvements have been achieved through the introduction of new therapeutic principles, such as the early addition of imatinib tyrosine kinase inhibitor with Ph-positive (Ph +) ALL (Lee et al., Blood 102 (2003), 3068-3070) or the use of anti-rituximab anti-CD20 antibody in ALL cases from CD20 + B-line cells (see, for example, Griffin et al., Pediatr Blood Cancer 2008). Diagnostic improvements were achieved by either using molecular genetic methods or flow cytometry to determine the level of minimum residual disease (MRD), which was found to be predictable for outcome in a number of studies in children (see, for example, Cave et al., N. Engl. J. Med. 339 (1998), 591-598) and adults (see, for example, Brüggemann et al., Blood 107 (2006), 1116-1123). Survival rates using modern treatment protocols for adult patients with acute lymphoblastic leukemia (ALL) have reached a plateau on which the possible benefits of more intensive chemotherapeutic regimens are often offset by excessive mortality due to complications, which makes, therefore, efforts to individualize treatment even more important. Whereas patients at ordinary risk are without usual risk factors whose chances of long-term survival using chemotherapy alone are greater than 50% (Hoelzer et al., Hematology Am. Soc. Hematol. Educ. Program 1 (2002), 162-192), at excessive risk with the intensification and prolongation of therapy, the outcome in patients with recurrent acute lymphoblastic leukemia (ALL) is very unfavorable, even if a second remission is achieved. In this study, the control of Minimum Residual Disease (MRD) during the first year of intensive chemotherapy entailed an MRD-based risk distribution (Bruggemann et al. (2006), cited). This classification allowed the establishment of a low-risk MRD group consisting of approximately 10% of patients with a minimum chance of relapse in 3 years, a high-risk MRD group consisting of approximately 25% of patients at almost 100% risk of relapse, and a medium-risk MRD group. In the latter group, approximately 30% of patients will eventually relapse despite becoming MRD negative or reaching MRD levels below 10 ^-4 at the end of the first year of therapy.

These data show that in most patients, acute lymphoblastic leukemia (ALL) remains a rapidly progressing and incurable disease. In view of this, there is an urgent need for improved therapies for ALL.

The present invention relates to a method for treating, reducing the intensity or elimination of acute lymphoblastic leukemia (ALL), comprising administering a pharmaceutical composition comprising the construction of a CD19 × CD3 bispecific single chain antibody to an adult patient in need thereof. In a preferred embodiment of the pharmaceutical methods and agents of the present invention, said acute lymphoblastic leukemia (ALL) is acute lymphoblastic leukemia (ALL) from B-line cells, preferably acute lymphoblastic leukemia (ALL) from B-cell progenitors. Acute lymphoblastic leukemia (ALL) from B-cell cells, accounting for 74% of cases, includes most ALL. Seventy percent of all ALL are ALL from precursor B cells, and 4% ALL are ALL from mature B cells. Since the CD19 × CD3 bispecific single chain antibody described herein is directed against the B19-linked CD19 marker, this antibody is particularly suitable as a therapeutic agent for acute lymphoblastic leukemia from B-line cells, preferably ALL from B-cells precursors that can be further subdivided into pro-B-cell ALL, pre-B-cell ALL and total B-cell ALL (c-ALL).

The introduction of a CD19 × CD3 bispecific single chain antibody (also called blinatumomab or MT103), described in more detail below, for the first time leads to a therapeutic approach that allows the treatment of minimal residual disease in patients with acute lymphoblastic leukemia (ALL). As shown in the following example and illustrated in FIG. 1, a CD19 × CD3 bispecific single chain antibody (the nucleic acid sequence and amino acid sequence of which is displayed in SEQ ID NO: 2 and 1, respectively) was designed to bind T cells to CD19 expressing target cells leading to unlimited cytotoxic T-cell response and activation of T-cells. Recently, a significant clinical activity of a CD19 × CD3 bispecific single chain antibody was demonstrated in a phase I test for recurrent B-cell non-Hodgkin's lymphoma (NHL) (Bargou et al., Science 321 (2008): 974-977). Based on these results, a phase II study was planned in conjunction with the German Multicenter Study Group on Adult Acute Lymphoblastic Leukemia (GMALL) to study the efficacy, safety, and tolerability of CD19 × CD3 bispecific single chain antibodies in patients with acute lymphoblastic leukemia (ALL) who achieved complete hematologic remission, but still had minimal residual disease (MRD). MRD is an independent prognostic factor that reflects resistance to the original drug (s) and is associated with a high risk of relapse after the onset of consolidation. MRD was determined using standard methods, either by quantifying individual rearrangements of immunoglobulin genes or rearrangements of T cell receptor (TCR) genes, translocations of t (4; 11), or the determination of bcr / abl fusion transcripts (see, for example, Van der Velden et al ., Leukemia 18 (2004), 1971-1980). The study population includes adult patients with acute lymphoblastic leukemia (ALL) from precursor B cells that show a signal for bcr / abl or a signal for translocation t (4; 11) that exceeds the detection limit and / or at least one marker for rearrangement with sensitivity ≥10 ^-4 . The initial expected result of a Phase II study is the degree of negative negative residual disease (MRD) status, determined by a signal with respect to bcr / abl or translocation t (4; 11), which is below the detection limit, and / or by identifying individual rearrangements of immunoglobulin genes or T-cell receptors (TCR) at a level that is below 10 ^-4 . One treatment cycle for a CD19 × CD3 bispecific single chain antibody is a 4-week continuous intravenous infusion, followed by transplantation of allogeneic hematopoietic stem cells after the first cycle or repeated cycles after a 2-week period without treatment. Dose CD19 × CD3-bispecific single chain antibody is 15 mcg / m ^2/24 hours, while allowing higher doses of one and the same patient, up to 30 microgram / m ^2/24 hours. The condition of minimum residual disease (MRD) is controlled after each treatment cycle. Patients who have reached MRD negativity may be given additional treatment cycles.

By this number, seventeen adult ALL patients have been treated, or are still being treated, with a CD19 × CD3 bispecific single chain antibody. 14 patients were assigned dose level is 15 mcg / m ^2/24 hours CD19 × CD3-bispecific single chain antibody, whereas three patients the dose was increased from 15 to 30 microgram / m ^2/24 hours after the first or further treatment cycles. All of these ALL patients had minimal residual disease (MRD): eleven of them had MRD determined by rearrangements of immunoglobulin or TCR genes, two patients had t translocations (4; 11), and four patients showed bcr / abl fusion transcripts.

As a result, the MRD response was measurable in 16 of 17 patients. 13 out of 16 patients became MRD-negative, which corresponds to an amazing degree of complete response at the molecular level of 81%. More specifically, MRD negativity could be achieved in nine of eleven patients with rearrangements of immunoglobulin or TCR genes, in one of two patients with t translocations (4; 11), and in three of four patients with bcr / abl transcripts. The largest period of MRD negativity observed so far in a patient who has not undergone transplantation after treatment with an antibody is 41 weeks. Another patient treated with a CD19 × CD3 bispecific single chain antibody that was MRD negative from 06/23/2008 to 10/27/2008 and who underwent successful transplantation of allogeneic hematopoietic stem cells after this, has not yet recurred.

It is noteworthy that patients with bcr / abl, who could be successfully treated with a CD19 × CD3 bispecific single chain antibody, in the previous ALL treatment regimen were resistant to imatinib and / or dasatinib tyrosine kinase inhibitors. For example, one of the bcr / abl patients who responded to treatment with a CD19 × CD3 bispecific single chain antibody had a T315I mutation that was resistant to tyrosine kinase inhibitor therapy. Thus, the introduction of a CD19 × CD3 bispecific single chain antibody is now the first to provide therapy for imatinib and / or dasatinib resistant patients with ALL with bcr / abl transcripts. Only three of the total 17 patients did not become MRD-negative. However, in two of them, stabilization of the disease could be achieved. Only one patient suffered a testicular relapse followed by hematologic relapse after 19 weeks of negativity in terms of the minimum residual disease. One patient could not be evaluated due to a serious side effect on day 2 of the study.

In conclusion, a definitely exceptional degree of complete response at the molecular level of 81% could be achieved in adult patients with ALL from precursor B cells as a result of treatment with a CD19 × CD3 bispecific single chain antibody. The activity of this antibody could be detected in all subgroups of patients treated with ALL, including those resistant to tyrosine kinase inhibitors (T315I) patients with bcr / abl and patients with translocations t (4; 11). These subgroups of patients with ALL were generally considered incurable using generally accepted standard therapy for ALL, with the exception of the allogeneic HSCT choice. In addition, when treating a CD19 × CD3 bispecific single chain antibody, a suitable toxicity profile is demonstrated, as opposed to conventional ALL therapies such as chemotherapy. In view of this, the administration of the CD19 × CD3 bispecific single chain antibody described herein provides a new and advantageous treatment option for acute lymphoblastic leukemia (ALL) in adults, especially in cases where ALL is resistant to conventional therapy for ALL, such as chemotherapy and / or allogeneic HSCT. In addition, the introduction of a CD19 × CD3 bispecific single chain antibody is now the first to determine therapy for MRD-positive ALL.

The method of the present invention provides the following main advantages:

1. Less adverse effects than conventional therapies for acute lymphoblastic leukemia (ALL), including chemotherapy or allogeneic HSCT. Conventional therapies for ALL are associated with significant health risks for patients; see, for example, Schmoll, Höffken, Possinger: Kompendium lnternistische Onkologie, S. 2660 ff .; 4. Auflage, Springer Medizin Verlag Heidelberg).

2. Although it is now believed that allogeneic HSCT is the preferred treatment for Ph + ALL, approximately one third of the transplanted patients suffer a relapse. Patients with Ph + ALL are most at risk of relapse among all patients within the ALL subtype. As shown in the following example, the administration of a CD19 × CD3 bispecific single chain antibody is particularly suitable for adult ALL sufferers with minimal residual disease (MRD). This is taking into account the minimum residual disease (MRD), determined by translocation with the formation of the Philadelphia chromosome, as well as MRD, determined by the rearrangement of immunoglobulin or TCR genes or translocation t (4; 11). Adult patients with ALL, not suitable for bone marrow transplantation, bearing t (4; 11), or resistant patients with Ph + ALL were still considered treatable. Therefore, the pharmaceutical methods and means of the present invention provide a therapeutic approach to treating, reducing the intensity or eliminating MRD ALL in adults, thereby reducing or even eliminating the risk of relapse for a patient. It should be noted that until now there has not been a cure for MRD-positive patients with ALL.

3. In particular, a CD19 × CD3 bispecific single chain antibody can be used to treat MRD-positive acute lymphoblastic leukemia (ALL) resistant to conventional therapies such as chemotherapy, administration of tyrosine kinase inhibitors and / or HSCT.

4. A CD19 × CD3 bispecific single chain antibody can not only replace conventional therapies for acute lymphoblastic leukemia (ALL) in patients not suitable for allogeneic HSCT, it can also be used to translate the condition of patients with ALL suitable for the indicated transplantation into MRD- negative state, since MRD-negative patients are at lower risk of relapse after transplantation than MRD-positive patients.

5. The high cytotoxic activity of the CD19 × CD3 bispecific single chain antibody allows the elimination of leukemia cells in the bone marrow.

A review of acute lymphoblastic leukemia (ALL), including acute lymphoblastic leukemia from precursor B cells and other types of ALL from B-line cells, and their treatments is provided, for example, in Pui and Evans, N. Engl. J. Med. 354 (2006), 166-178; Hoelzer and Gökbuget; Hematology (2006); 133-141; or Apostolidou et al., Drugs 67 (2007), 2153-2171. Information regarding ALL can also be found, for example, at http://www.cancer.gov, http://www.wikipedia.org or http://www.leukemia-lymphoma.org.

The term “bispecific single chain antibody” or “single chain bispecific antibody” or related terms in accordance with the present invention means antibody constructs resulting from the joining of at least two variable regions of antibodies into a single polypeptide chain free of constant regions and / or Fc- part (s) present in full-sized immunoglobulins. A linker connects V domains of the same specificity, while a spacer connects V domains of different specificities. For example, a bispecific single chain antibody can be a construct in which only two variable regions of antibodies are used, for example, two VH regions, each of which is capable of specific binding of a particular antigen, and which are connected to each other via a short (usually less than 10 amino acids) synthetic a polypeptide spacer so that two variable regions of antibodies with a spacer inserted between them exist as a single continuous polypeptide chain. Another example of a bispecific single chain antibody may be a single polypeptide chain with three variable regions of antibodies. In this case, two variable regions of antibodies, for example, one VH and one VL, can be scFv, in which two variable regions of the antibodies are connected to each other via a synthetic polypeptide linker, the linker is often genetically designed so that it is minimally immunogenic when maintaining maximum resistance to proteolysis. This scFv is capable of specific binding to a particular antigen and is coupled to an additional variable region of the antibody, for example a VH region capable of binding to an antigen other than that which binds to scFv. Another example of a bispecific single chain antibody can be a single polypeptide chain with four variable regions of antibodies. In this case, the first two variable regions of antibodies, for example, the VH region and the VL region, can form one scFv capable of binding to one antigen, while the second two variable regions: VH and VL - can form a second scFv capable of binding to another antigen. Within a single continuous polypeptide chain, the individual variable regions of antibodies of the same specificity can be predominantly separated by the synthetic polypeptide linker described above, while the corresponding scFv can be predominantly separated by the short polypeptide spacer described above. Non-limiting examples of bispecific single chain antibodies, as well as methods for their preparation, are presented in WO 99/54440, WO 2004/106381, WO 2007/068354, Mack, J. Immunol. (1997), 158, 3965-3970; Mack, PNAS, (1995), 92, 7021-7025; Kufer, Cancer Immunol. Immunother, (1997), 45, 193-197; Loffler, Blood, (2000), 95, 6, 2098-2103; Bruhl, J. Immunol., (2001), 166, 2420-2426.

"CD3" refers to an antigen that is expressed on T cells, preferably human T cells, as part of a multi-molecular T-cell receptor complex, wherein CD3 consists of five different chains: CD3 epsilon, CD3 gamma, CD3 delta, CD3 -eta and CD3-zeta. Aggregation of CD3 on T cells, for example, with anti-CD3 antibodies, results in T-cell activation similar to that of antigen binding, but independent of the clonal specificity of the T-cell subpopulation. Thus, the term “CD19 × CD3 bispecific single chain antibody” refers to a CD3-specific construct capable of binding to a human CD3 complex expressed on human T cells and capable of inducing elimination / lysis of target cells, such cells the targets carry / represent an antigen that binds to the other non-CD3 binding part of the bispecific single chain antibody. The binding of a CD3 comprising complex to CD3-specific binding agents (for example, a bispecific single chain antibody administered in accordance with the pharmaceutical agents and methods of the present invention) results in T-cell activation, as is known in the art; see, for example, WO 99/54440 or WO 2007/068354. Accordingly, a construct suitable for the pharmaceuticals and methods of the present invention is preferably capable of eliminating / lysing target cells in vivo and / or in vitro. Suitable target cells include cells expressing a tumor antigen, such as CD19, which is recognized by the second specificity (i.e., the non-CD3 binding portion of the bispecific single chain antibody) of the indicated construct. Preferably, said second specificity is specificity for human CD19, which has already been described in WO 99/54440, WO 2004/106381 or WO 2007/068354. In accordance with this embodiment, each antigen-specific portion of a bispecific single chain antibody comprises an antibody VH region and an antibody VL region. An advantageous variant of this bispecific single chain antibody is the following variant in the direction from the N-terminus to the C-terminus:

V _L (CD19) -V _H (CD19) -V _H (CD3) -V _L (CD3) (SEQ ID NO: 1).

In the meaning of the present invention, the term “specifically binding” or related terms such as “specificity” should be understood to be characterized mainly by two parameters: a qualitative parameter (binding epitope, or where the antibody binds) and a quantitative parameter (binding affinity, or how strongly this antibody binds where it binds). Which epitope is bound by an antibody can advantageously be determined, for example, using a technique using FACS, ELISA, epitope mapping using peptide regions or mass spectroscopy. The binding strength of an antibody to a particular epitope can be advantageously determined, for example, using known Biacore and / or ELISA techniques. The combination of these methods allows us to calculate the signal: noise ratio as a representative indicator of the specificity of binding. In this respect, signal: noise, the signal represents the binding force of the antibody to the epitope of interest, while the noise represents the binding force of the antibody to other, unrelated epitopes other than the epitope of interest. The signal-to-noise ratio, for example, of at least 50, but preferably approximately 80, for the corresponding epitope of interest, as determined, for example, by Biacore, ELISA or FACS, can be taken as an indicator that the antibody being evaluated is bound with the epitope of interest in a specific way, i.e. it is a “specific binder”. The term “binding to / interacting with” may also refer to a conformational epitope, a structural epitope, or a discontinuous type determinant consisting of two or even more regions of human target molecules or parts thereof. The conformational epitope is defined by two or more discrete amino acid sequences, separated in the primary sequence, which converge on the surface of the molecule when the polypeptide is packed into a naturally occurring protein (Sela, (1969) Science 166, 1365 and Laver, (1990) Cell 61, 553-556 ) The term “discontinuous type determinant” means non-linear epitopes that are assembled from residues from spaced portions of a polypeptide chain. These residues converge on the surface of the molecule when the polypeptide chain is laid in a three-dimensional structure with the creation of a conformational / structural epitope.

In its broadest sense, the term "treatment" means medical procedures or applications that are intended to alleviate the disease. In the case of the present invention, the administration of a CD19 × CD3 bispecific single chain antibody (prepared for administration to an adult patient with ALL) described herein is performed to treat, reduce the intensity or eliminate the ALL disease in adult patients.

The term “patient” refers to an adult patient. As used herein, the term “ALL in adults” or “adult patient with ALL” or “adult patient” means adults over the age of 18 years, i.e. patients aged 19, 20, 21, 22, 23, 24, 25, 30, 35, 40, or 50 years or older. Patients age 70, 75, 80, 85, 90, 100, 100 years or older may even be treated using the methods and means of the present invention. It should be understood that this age is the age of adults in diagnosing ALL disease.

The term “intensity reduction” is synonymous with the term “improvement”. If the condition of an adult patient with ALL shows improvement, the patient is obviously better - there is some improvement in her or his condition. For example, in the condition of a patient with ALL, there may be an improvement if stabilization of ALL disease (stage of the disease, also called stable disease) can be achieved, i.e. ALL disease is no longer progressive. Even better, when MRD-positive acute lymphoblastic leukemia (ALL) translates into an MRD-negative state.

The term "elimination (elimination)" means the removal of leukemia cells from the body of an adult patient with ALL. As shown in the following example, the administration of a CD19 × CD3 bispecific single chain antibody is capable of translating MRD-positive acute lymphoblastic leukemia (ALL) into an MRD-negative state for various ALL subtypes.

The term “administration” means the administration of a therapeutically effective dose of the aforementioned CD19 × CD3 bispecific single chain antibody to an individual, i.e. a human patient. Preferably, the ALL patient is an adult patient as defined herein.

By “therapeutically effective amount” is meant a dose that causes the effects for which it is administered, preferably the conversion of the MRD-positive state of acute lymphoblastic leukemia (ALL) to the MRD-negative state ALL. The exact dose will depend on the purpose of the treatment and can be determined by a person skilled in the art using known methods. As is known in the art and described above, harmonization with systemic delivery may be required compared with local delivery, age, body weight, general health, gender, diet, management time, drug interactions and severity, and such matching can be established by those skilled in the art using routine experimentation.

The staff doctor and clinical factors will determine the dosage regimen. As is well known in the medical field, doses for any one patient depend on many factors, including size, body surface area, age, patient body weight, specific compound administered, gender, time and route of administration, general health status, and other medicinal products funds administered simultaneously.

As is well known in the medical field, doses for any one patient depend on many factors, including size, body surface area, age, patient body weight, specific compound administered, gender, time and route of administration, general health status, and other medicinal products funds administered simultaneously. A typical dose may, for example, be in the ranges set forth in embodiments of the present invention and the accompanying examples; however, doses are provided that are lower or higher than this exemplary range, especially taking into account the above factors.

The term “continuous infusion” refers to an infusion that is allowed to be administered continuously over a period of time, i.e. nonstop. “Continuous infusion” refers to a continuous infusion. Accordingly, in the context of the present invention, the terms “constant” and “continuous” are used as synonyms. For the purposes of the present invention, for example, the term “continuous infusion over 4 weeks” refers to a situation in which a CD19 × CD3 bispecific single chain antibody used in the pharmaceutical agents and methods of the present invention is continuously administered to an adult patient for a period of constituting 4 weeks, in a continuous, constant manner throughout the entire period required in the pharmaceutical products and methods of the present invention. Schemes for continuous administration of a CD19 × CD3 bispecific single chain antibody are described in more detail in WO 2007/068354. Avoid interruption in the administration of a CD19 × CD3 bispecific single chain antibody, i.e. the transition from a situation in which an antibody is administered to a patient to a situation in which this antibody is no longer administered to a patient during the entire administration period required by pharmaceutical agents and methods does not take place, or substantially does not take place of the present invention, for reasons other than replenishment of the administered CD19 × CD3 bispecific single chain antibody or medical interventions that have become necessary, etc. Since such necessary replenishment leads to a temporary interruption in the administration of the administered antibody, such administration should still be considered “uninterrupted” or “permanent” in the meaning of the pharmaceutical agents and methods in accordance with the present invention. In most cases, usually the duration of such replenishment will be so short that the time during which the antibody is not introduced into the patient’s body will be vanishingly small compared to the time planned for the entire administration schedule in accordance with the pharmaceutical agents and methods of the present invention. In accordance with the present invention, a single cycle of administration should be understood as continuous infusion of a CD19 × CD3 bispecific single chain antibody to an adult patient with ALL for a period of 4 weeks, followed by a 2-week period without treatment. It is possible that after determining the stage of MRD in the treated patient (s) after continuous administration for 4 weeks or one treatment cycle, a minimal or partial response to treatment with a bispecific single chain antibody can be diagnosed. In this case, continuous administration can be continued for an additional one, two, three, four, five, or even up to ten treatment cycles to achieve a better therapeutic result, for example, stabilization of the disease or even a complete response. Preferably, said full response is MRD negativity. In an alternative embodiment, continuous infusion of a CD19 × CD3 bispecific single chain antibody into an adult ALL patient for 4 weeks may be followed by allogeneic HSCT. It is also contemplated that a patient subjected to treatment using one, two, three, four or even more treatment cycles, as described above, can then be transplanted with allogeneic hematopoietic stem cells.

As shown in the following example, as a result of treatment with CD19 × CD3 bispecific single chain antibody, 13 of 16 adult ALL patients became MRD negative, which corresponds to an amazing degree of complete response at the molecular level of 81%. More specifically, negativity in terms of the minimum residual disease could be achieved in nine of eleven patients with rearrangements of immunoglobulin or TCR genes, in one of two patients with t translocations (4; 11), and in three of four patients with bcr / abl transcripts. Preferably, the main therapeutic goal of administering a CD19 × CD3 bispecific single chain antibody, either alone or in combination with allogeneic HSCT, to an adult ALL patient is to convert the MRD-positive state to the MRD-negative state as defined herein.

Continued continuous administration of the bispecific single chain antibody in the style of pharmaceuticals and methods in accordance with the present invention for longer periods of time makes it possible to preferentially activate the T cells indicated in the examples with a manifestation of its effect for a long time, sufficient for the preferred removal of all diseased cell disease from the body. Since the rate of continuous administration of a bispecific single chain antibody is kept low, the use of a therapeutic agent can last longer without risking harmful side effects for the patient.

Preferably, the CD19 × CD3 bispecific single chain antibody used is in the form of a pharmaceutical composition for administration to a human patient diagnosed with acute lymphoblastic leukemia (ALL). Preferably, the human patient is an adult, as defined below. Although the bispecific single chain antibody used herein can be administered alone, administration in a pharmaceutically acceptable carrier is preferred. Examples of suitable pharmaceutically acceptable carriers are well known in the art and include phosphate buffered saline, water, liposomes, various types of wetting agents, sterile solutions, etc. Compositions comprising such carriers can be formulated using well-known conventional methods. These pharmaceutical compositions may be administered to a subject in an appropriate dose. The staff doctor and clinical factors will determine the dosage regimen. As is well known in the medical field, doses for any one patient depend on many factors, including size, body surface area, age, patient body weight, specific compound administered, gender, time and route of administration, general health status, and other medicinal products funds administered simultaneously. Preparations for parenteral administration include sterile aqueous or non-aqueous solutions or suspensions. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, and injectable organic acid esters such as ethyl oleate. Aqueous carriers include water, aqueous solutions or suspensions, including saline and buffered media. Carriers for parenteral administration include sodium chloride solution, dextrose in Ringer's solution, dextrose and sodium chloride, or Ringer's solution with lactate. Carriers for intravenous administration include liquid and food additives, electrolyte additives (such as those based on dextrose in Ringer's solution) and the like. Preservatives and other additives may also be present, such as, for example, antimicrobials, antioxidants, chelating agents and inert gases and the like. In addition, the composition could include protein-like carriers, such as, for example, serum albumin or immunoglobulin, preferably of human origin. It is possible to include, in addition to a protein-like bispecific single chain antibody, additional biologically active agents, depending on the intended use of the pharmaceutical composition. Such agents could be agents acting as cytostatics, agents for preventing hyperuricemia, agents that inhibit immune reactions (e.g., corticosteroids, FK506), drugs that have an effect on the cardiovascular system, and / or agents such as costimulatory T β-cells of a molecule or cytokines known in the art.

Preferably, a mixture of the CD19 × CD3 bispecific single chain antibody as defined herein with a buffer, stabilizer and surfactant is formulated. The buffer may be phosphate, citrate, succinate or acetate buffer. The stabilizer may be amino acid (s) and / or sugar. Surfactants may be detergents, PEG, or the like. More preferably, when a mixture of a CD19 × CD3 bispecific single chain antibody, as defined herein, is made up of citrate, lysine, trehalose and Tween 80. Isotonic saline and Tween 80 are preferred as diluent for the pharmaceutical composition of the present invention.

Preferably, in the uses or methods of the present invention, the pharmaceutical composition should be administered to a human adult patient diagnosed with acute lymphoblastic leukemia (ALL).

The viability of therapy using a CD19 × CD3 bispecific single chain antibody can be controlled using well-known standard methods for the respective disease categories: in the case of therapy for B-cell ALL, analysis using a fluorescence-excited cell sorter (FACS), bone marrow aspiration and various specific for leukemia, clinical chemical parameters, and other well-known standard methods. Methods and means for determining the state of MRD are described herein.

Cytotoxicity can be determined using methods known in the art and methods illustrated, for example, in WO 99/54440, WO 2004/106381, WO 2007/068354.

In a preferred embodiment, chemotherapy resistant is acute lymphoblastic leukemia (ALL) of an adult patient (s), preferably in terms of MRD (i.e., MRD in these ALL patients is chemotherapy resistant). Even more preferably, acute lymphoblastic leukemia (ALL) is chemotherapy resistant in patients not suitable for allogeneic stem cell transplantation.

The term “chemotherapy” refers to chemotherapy used to treat acute lymphoblastic leukemia (ALL). Chemotherapy is the initial preferred treatment for ALL. Most patients with ALL find themselves receiving a combination of different treatments. In the treatment of ALL, there is no choice of surgery due to the spread of malignant cells throughout the body. Typically, cytotoxic chemotherapy for ALL combines many anti-leukemia drugs in various combinations. Chemotherapy for ALL consists of three steps: inducing remission, intensification, and supportive care. Chemotherapy is also prescribed to protect the central nervous system from leukemia. The goal of inducing remission is the rapid destruction of most tumor cells and the introduction of a patient into remission. It is defined as the presence of less than 5% leukemic blast cells in the bone marrow (as determined by optical microscopy), normal blood cells and the absence of tumor cells in the blood, and the absence of other signs or symptoms of the disease. To induce remission, for example, a combination of prednisone or dexamethasone (in children), vincristine, asparaginase and daunorubicin (used in ALL in adults) is used. Intensification uses high doses of chemotherapy using a variety of intravenous drugs to further reduce the tumor mass. Typical intensification protocols use vincristine, cyclophosphamide, cytarabine, daunorubicin, etoposide, thioguanine, or mercaptopurine, which are assigned as groups in various combinations. Since sometimes ALL cells enter the central nervous system (CNS), most protocols include delivering chemotherapy to the cerebrospinal fluid (called intrathecal chemotherapy). Some centers deliver the drug thanks to the Ommaya reservoir (a device that is surgically placed under the scalp and used to deliver drugs to the cerebrospinal fluid and to extract cerebrospinal fluid for various tests). Other centers make many lumbar punctures, as necessary, for analysis and treatment delivery. Intrathecal methotrexate or cytarabine is usually used for this purpose. The goal of maintenance therapy is to destroy any surviving cell that was not destroyed during the remission and intensification schemes. Although there are few such cells, they will cause a relapse if they are not destroyed. For this purpose, mercaptopurine is used daily, given orally, once a week, methotrexate, given orally, once a month, a 5-day course of intravenous administration of vincristine and orally prescribed corticosteroids. The duration of maintenance therapy is 3 years for boys, 2 years for girls and adults. Recurrences of the central nervous system lesions treated with intrathecal administration of hydrocortisone, methotrexate, and cytarabine (Hoffbrand et al., Essential Hematology , Blackwell, 5 ^th edition, 2006). Since chemotherapy regimens can be intense and lengthy (often for about 2 years for GMALL UKALL, HyperCVAD or CALGB protocols; for about 3 years for men in COG protocols), many patients have an intravenous catheter inserted into the central vein (called a central venous or Hickman catheter), or Portacath (a cone-shaped small device with a silicone nose that is surgically inserted under the skin, usually around the collarbone).

Chemotherapy for ALL is described, for example, in Schmoll, Hoffken, Possinger (in the cited place).

In view of the foregoing, the term “chemotherapy resistant” as used herein refers to the resistance of chemotherapy to acute lymphoblastic leukemia cells.

Patients may tolerate ALL relapse after initial therapy and / or become chemotherapy resistant after treatment. Chemotherapy-resistant patients with ALL have a clearly poor prognosis. In particular, the prognosis is unfavorable for adult patients with Ph + ALL who underwent only treatment with chemotherapy, while the probability of prolonged survival is less than 10%. Because the pharmaceutical methods and agents of the present invention can lead to MRD negativity in adult ALL patients, they are particularly suitable for treating chemotherapy resistant ALL patients.

The term "allogeneic hematopoietic stem cell transplantation" means allogeneic hematopoietic stem cell transplantation (HSCT) or bone marrow transplantation (BMT), which is a medical procedure in the field of hematology and oncology, which includes hematopoietic stem cell transplantation (HSC). Most often, it is performed in the case of patients with diseases of the lymph nodes, blood or bone marrow, or some types of cancer, such as ALL. Allogeneic HSCT is a procedure in which a person receives hematopoietic stem cells (cells from which all blood cells are formed) from a genetically similar but not identical donor. It is often a close relative, such as a mother, father, sister or brother, but could also be a non-relative donor. Most HSCT recipients are patients with leukemia (e.g. ALL) who might benefit from treatment with high doses of chemotherapy or whole body irradiation. However, allogeneic HSCT remains a dangerous and toxic treatment.

The term “not suitable for HSCT” is intended for those adult patients in whom allogeneic HSCT is not the preferred treatment for ALL, for example, for medical reasons. For example, it is possible that there is no suitable donor available or the patient has already exceeded the upper age limit. As shown in the following example, all patients were resistant to chemotherapy, or in the case of Ph + ALL, also resistant to tyrosine kinase inhibitors or not tolerating them before inclusion in the study. Eight patients treated with CD19 × CD3 bispecific single chain antibody, such as, for example, patients 111-003, 108-002, 109-006 or 109-007, were not suitable for allogeneic HSCT.

To date, ALL has meant sentencing for patients who are resistant to chemotherapy and are not suitable for allogeneic HSCT. The pharmaceutical methods and means of the present invention for the first time provide treatment for this patient population, which eliminates the minimum residual disease (MRD), which otherwise could cause a relapse and kill these patients.

In an alternative embodiment of the pharmaceutical methods and agents of the present invention, said method is followed by transplantation of allogeneic hematopoietic stem cells, or the method replaces transplantation of allogeneic hematopoietic stem cells in adult patients suitable for allogeneic HSCT.

The term “suitable for allogeneic HSCT” means that allogeneic HSCT is a necessary therapy for an adult patient with ALL. In cases where a patient with ALL is suitable for allogeneic HSCT, the following two scenarios can be considered. First, in one embodiment of the pharmaceutical methods and means of the present invention, the administration of a CD19 × CD3 bispecific single chain antibody (alone or preferably in the form of a pharmaceutical composition) can be used to replace allogeneic HSCT used as a standard therapy for adult ALL patients suitable for transplantation. Therefore, the pharmaceutical methods and means of the present invention can prevent the health risks of patients with ALL associated with transplantation of allogeneic hematopoietic stem cells. In addition, approximately 30% of transplanted patients with ALL usually suffer a relapse after transplantation. Therefore, the pharmaceutical methods and agents of the present invention can be used to treat these patients. In an alternative embodiment, continuous infusion of a CD19 × CD3 bispecific single chain antibody into an adult ALL patient may be followed by allogeneic stem cell transplantation. In this embodiment, the administration of a pharmaceutical composition comprising the construction of a CD19 × CD3 bispecific single chain antibody can be used to translate the condition of patients with ALL suitable for transplantation into an MRD negative state prior to transplantation. Therefore, the pharmaceutical methods and means of the present invention can be used to eliminate MRD, which leads to a lower risk of relapse than treatment using transplantation of MRD-positive patients. The example shows a patient who was first transferred to an MRD negative state as a result of treatment with a CD19 × CD3 bispecific single chain antibody, followed by allogeneic transplantation. So far, this patient is still MRD-negative, with a period of MRD-negativity of 47 weeks to date.

Within the scope of the pharmaceutical methods and means of the present invention is also the introduction of the construction of a CD19 × CD3 bispecific single chain antibody to adult ALL patients who have undergone allogeneic HSCT and suffered a relapse thereafter.

In another preferred embodiment, the pharmaceutical methods and agents of the present invention are for treating, reducing the intensity or eliminating minimum residual disease (MRD) in an adult patient with acute lymphoblastic leukemia (ALL).

The term "minimum residual disease (MRD)" refers to the condition of the disease after treatment with, for example, chemotherapeutic agents, when leukemia cells can no longer be detected in the bone marrow using methods using optical microscopy. More sensitive assays, such as flow cytometry (FACS based methods) or polymer chain reaction (PCR), should be used to find evidence that leukemia cells remained in the bone marrow of a patient with ALL. More specifically, the presence of leukemic cells at a level that is below the cytological detection limit (5% of leukemic cells) is defined as “minimum residual disease (MRD)”. If MRD is not detected (<10 ^-4 , i.e., less than 1 leukemia cell per 10 ⁴ detectable bone marrow cells), complete remission at the molecular level is achieved. By “MRD-positive state,” as defined herein, is meant a signal for bcr / abl or a signal for translocation t (4; 11) that exceeds the detection limit and / or at least one marker for individual immunoglobulin gene rearrangements or T cell receptors (TCR), in excess of 10 ^-4 . By “MRD negative state”, as defined herein, is meant a signal for bcr / abl or a signal for translocation t (4; 11) that is below the detection limit, or at least one marker for individual gene rearrangements of immunoglobulins or T cell receptors (TCR), at a level that is below 10 ^-4 . MRD status can be determined using PCR or FACS analysis, which quantifies individual rearrangements of immunoglobulin genes or rearrangements of T-cell receptors (TCR), or transcripts of bcr / abl fusion, or translocation t (4; 11). For example, using PCR analysis, fusion transcripts, such as bcr / abl, or translocations of t (4; 11), as well as individual clonal rearrangements of immunoglobulin (IgH) and / or T-cell receptor (TCR) genes, can be detected.

Repeated chromosomal abnormalities in the malignant cells of patients with acute lymphoblastic leukemia are signs of the disease (Harrison and Foroni, Rev. Clin. Exp. Hematol. 6 (2002), 91-113). Specific aberrations, which often serve as a sign of regular, underlying damage at the molecular level, can help diagnose or even establish a diagnosis and determine the optimal therapy. In the case of ALL in children, numerous cytogenetic subgroups of low and high risk have been identified that are regularly used to refer patients to specific therapies (Pui and Evans, N. Engl. J. Med. 354 (2006), 166-178). However, in the case of ALL in adults, the role of cytogenetics in the treatment of patients was mainly focused on the presence of the Philadelphia (Ph) chromosome, which usually occurs as a result of translocation t (9; 22) (q34; q11.2) and leads to the fusion of BCR- ABL (bcr / abl) (Faderl et al., Blood 91 (1998), 3995-4019). Although the overall incidence of Ph + ALL in adults is approximately 25%, it is associated with age and increases to over 50% among patients over 55 years old (Appelbaum, American Society of Clinical Oncology 2005 education book. Alexandria: ASCO, 2005: 528 -532). Other cytogenetic translocations associated with specific molecular genetic abnormalities in acute lymphoblastic leukemia (ALL) are presented in Table 1.

Table 1 Cytogenetic translocation Molecular genetic deviation t (9; 22) (q34; q11) BCR-ABL Merger (P185) t (12; 21) CRYPTIC TEL-AML1 merger t (1; 19) (q23; p13) E2A-PBX Merge t (4; 11) (q21; q23) MLL-AF4 merger t (8; 14) (q24; q32) IGH-MYC merger t (11; 14) (p13; q11) TCR-RBTN2 merger

Cytogenetics has been increasingly recognized as an important prognostic factor in the outcome of ALL (Moormann et al., Blood 109 (2007), 3189-3197).

Some cytogenetic subtypes have a poorer prognosis than others. These, for example, include:

(i) Translocation between chromosomes 9 and 22, the Philadelphia chromosome (Ph +), occurs in approximately 20% of adults and 5% of pediatric patients with ALL.

(ii) Translocation between chromosomes 4 and 11 occurs in approximately 4% of patients and most often in infants less than 12 months old.

Rearrangement of immunoglobulin genes or rearrangement of T-cell receptor (TCR) genes and their role in ALL are described in the art (see, for example, Szczepanski et al., Leukemia 12 (1998), 1081-1088).

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, said adult patient is MRD positive with complete hematologic remission.

The term "remission" or "hematological remission" should be understood as the absence of data on the disease after treatment, for example, after chemotherapy or transplantation. This means that the bone marrow contains less than 5% of blast cells determined by optical microscopy, blood cell levels are within normal limits, and there are no signs or symptoms of ALL disease. “Complete remission at the molecular level” means the lack of data on leukemia cells in bone marrow biopsies, even after using very sensitive assays, such as PCR. In other words, if MRD is not detected (<10 ^-4 , i.e., less than one leukemia cell per 10 ⁴ bone marrow cells), complete remission at the molecular level is achieved.

After complete remission of the lesion (s) by leukemia in a human adult patient with ALL achieved with chemotherapeutic treatment or transplantation of allogeneic hematopoietic stem cells, it is possible that it was impossible to eliminate all leukemic cells from the body. However, these remaining tumor cells can cause the development of recurrent leukemia. The pharmaceutical agents and methods of the present invention can be used to kill these remaining tumor cells in order to prevent the recurrence of leukemia (arising from undetectable leukemia cells remaining in the body after the initial therapy). Thus, pharmaceutical agents and methods help prevent relapse in adult patients with ALL.

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, administering said pharmaceutical composition converts MRD-positive acute lymphoblastic leukemia (ALL) into an MRD-negative state.

In another preferred embodiment of the pharmaceutical methods and means of the present invention, MRDs are determined by quantifying individual rearrangements of immunoglobulin genes or rearrangements of T cell receptor (TCR) genes, or bcr / abl fusion transcripts, or t translocation (4; 11) using analysis using PCR or FACS.

As shown in the following example, the administration of a CD19 × CD3 bispecific single chain antibody is particularly suitable for adult patients with minimal residual disease (MRD). This is taking into account the minimum residual disease (MRD), determined by translocation with the formation of the Philadelphia chromosome or translocation t (4; 11), as well as MRD, determined by the rearrangement of immunoglobulin or TCR genes. The pharmaceutical methods and means of the present invention provide a therapeutic approach to treating, reducing the intensity or eliminating MRD, thereby reducing or even eliminating, thereby eliminating the risk of relapse for an adult patient. It is noteworthy that, until now, there was no curable MRD treatment in patients with ALL.

In another preferred embodiment of the pharmaceutical methods and means of the present invention, said patient exhibits a signal for bcr / abl or a signal for translocation t (4; 11) that exceeds the detection limit and / or at least one marker for rearrangement with sensitivity ≥10 ^-4 .

The term “bcr / abl signal or translocation signal t (4; 11) that exceeds the detection limit” means that analysis using PCR or FACS results in a detectable signal for bcr / abl or a signal for translocation t (4; 11).

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, the period of time before relapse at the molecular level (determined using the assays described above) is more than 4 months.

The term “relapse at the molecular level” means that the indicated patient shows a signal for bcr / abl or translocation t (4; 11) that exceeds the detection limit and / or at least one marker for rearrangement with sensitivity ≥10 ^-4 .

The term “with a sensitivity of ≥10 ^-4 ” means that one or more of one leukemic cell can be detected among 10,000 cells, in particular bone marrow cells.

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, the corresponding variable regions of the heavy chains (V _H ) and the corresponding variable regions of the light chains (V _L ) in the indicated construction of the CD19 × CD3 bispecific single chain antibody are located, from the N-terminus to C- end, in the following order, V _L (CD19) -V _H (CD19) -V _H (CD3) -V _L (CD3).

The corresponding variable regions of the heavy chains (V _H ) and the corresponding variable regions of the light chains (V _L ) of the binding CD3 and CD19 domains of the CD19 × CD3 bispecific single chain antibody are presented in SEQ ID NO: 3-10, respectively. The corresponding CDR regions of the corresponding VH and VL regions of the indicated CD19 × CD3 bispecific single chain antibody are presented in SEQ ID NO: 11-22.

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, said CD19 × CD3 bispecific single chain antibody construct comprises at least 90% the amino acid sequence reflected in SEQ ID NO: 1 or the amino acid sequence identical to SEQ ID NO: 1, preferably at least 95%.

The present invention describes a bispecific single chain antibody molecule comprising the amino acid sequence shown in SEQ ID NO: 1, as well as an amino acid sequence identical to the amino acid sequence of SEQ ID NO: 1 by at least 90% or preferably 95%, most preferably at least at least 96, 97, 98 or 99%. The present invention also describes the corresponding nucleic acid sequence shown in SEQ ID NO: 2, as well as a nucleic acid sequence identical to the nucleic acid sequence shown in SEQ ID NO: 2, at least 90%, preferably 95%, most preferably at least 96, 97, 98, or 99%. It should be noted that sequence identity is determined throughout the nucleotide or amino acid sequence. In addition, it should be noted that a bispecific single chain antibody molecule comprising an amino acid sequence identical to the amino acid sequence of SEQ ID NO: 1 is at least 90% or preferably 95%, most preferably at least 96, 97, 98 or 99 % contains all of the CDR sequences shown in SEQ ID NO: 11-22. For sequence matching, for example, Gap or BestFit programs can be used (Needleman and Wunsch J. Mol. Biol. 48 (1970), 443-453; Smith and Waterman, Adv. Appl. Math 2 (1981), 482-489), contained in the GCG software package (Genetics Computer Group, 575 Science Drive, Madison, Wisconsin, USA 53711 (1991). This combination is common for those skilled in the art to determine and identify a nucleotide or amino acid sequence identical to the nucleotide or amino acid sequence CD19 × CD3 bispecific single chain antibody, describing in the present description, for example, 90%, 95%, 96%, 97%, 98% or 99%. For example, in accordance with the hypothesis of ambiguous correspondence of Scream 5 ', the base in the anticodon is not as spatially limited as the other two bases and could therefore have non-standard base pairing, in other words, the third position in the triplet may be different, so two triplets that differ in this third position can encode the same amino acid residue. This hypothesis is well known to those skilled in the art (see, for example, http://en.wikipedia.org/wiki/Wobble_Hypothesis; Crick, J. Mol. Biol. 19 (1966): 548-555).

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, one treatment cycle is a continuous infusion for 4 weeks, followed by repeated cycles after a 2-week period without treatment or followed by transplantation of allogeneic hematopoietic stem cells.

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, the treatment cycle is repeated at least three times, preferably four, five, six, seven, or even up to ten times, after determining the MRD negative state (consolidation).

In another preferred embodiment of the pharmaceutical methods and agents of the present invention, the bispecific single chain antibody construct is to be administered in a daily dose of 10 μg to 100 μg per square meter of patient surface area.

As used herein, a dose range that is defined as “X to Y” is equivalent to a dose range that is defined as “between X and Y”. The range includes an upper limit as well as a lower limit. This means that, for example, a daily dose of 10 μg to 100 μg per square meter of patient’s body surface area includes “10 μg” and “100 μg”.

In an even more preferred embodiment of the pharmaceutical methods and agents of the present invention, a CD19 × CD3 bispecific single chain antibody construct should be administered at a daily dose of 15 μg, 30 μg, 60 μg or 90 μg per square meter of patient surface area. Even more preferably, said antibody should be administered in a daily dose of 15 μg to 30 μg per square meter of patient surface area, most preferably in a daily dose of 15 μg to 30 μg per square meter of patient surface area .

The average body surface area of an adult patient, as calculated, therefore, in relation to the pharmaceutical method or use in accordance with the present invention, is in the range from about 1.7 to 2.2 m ² .

Preferably, the pharmaceutical composition comprising the CD19 × CD3 bispecific single chain antibody described herein further comprises optionally working buffer solution (s), storage solutions (stock solutions) and / or other reagents and materials necessary for the method described above or application. In addition, these components can be packaged individually in bottles or bottles, or in combination in containers or multi-container units.

In order to evaluate the safety and tolerability of the CD19 × CD3 bispecific single chain antibody described herein, the compound must be administered by continuous continuous infusion.

It has been found that the positive and unexpected effects of the pharmaceutical agents and methods of the present invention can be achieved by administering a CD19 × CD3 bispecific single chain antibody in a daily dose of 10 μg to 100 μg per square meter of body surface area. The daily dose may remain constant during the administration period. However, it is also within the scope of this embodiment that a lower dose of a bispecific single chain antibody (“initial dose”) is administered during the initial day (s) of the infusion period before performing the pharmaceutical methods described herein, while during the rest of the infusion period, higher dose ("maintenance dose"). For example, you can enter 5 micrograms of a bispecific single chain antibody per square meter of body surface area on the first day (s) of the infusion period, followed by 15 micrograms per square meter of body surface area as a daily dose for the rest of the treatment period. Or, 15 micrograms of a bispecific single chain antibody per square meter of body surface area can be administered on the first day (s) of the infusion period, followed by 30 or 45 micrograms per square meter of body surface area as a daily dose for the rest of the treatment period. The initial dose may be administered within one, two or more days, or even within one week (seven days). It is also contemplated that 5 micrograms of a bispecific single chain antibody per square meter of body surface area can be administered on the first day (s) of the infusion period, followed by the administration of 15 micrograms of a bispecific single chain antibody per square meter of body surface area for the next day (days) of the infusion period and then inject 45 micrograms per square meter of body surface area as a daily (maintenance) dose for the rest of the treatment period. The average body surface area of an adult patient, as calculated, therefore, in relation to pharmaceutical methods or applications in accordance with the present invention, is in the range from 1.7 to 2.2 m ² .

In another embodiment, the methods and uses of the present invention the dosage is increased after the first or further treatment cycles, for example, from 15 to 30 or even 60 or 90 micrograms / m ^2/24 hours.

The continuous administration of a CD19 × CD3 bispecific single chain antibody can be intravenous, parenteral, subcutaneous, transdermal, intraperitoneal, intramuscular or pulmonary. In most cases, an intravenous route of administration will be the preferred method for continuous administration of a CD19 × CD3 bispecific single chain antibody and, as the case may be, for co-administration of a pharmaceutical agent as part of a co-therapy regimen. As such, intravenous administration is particularly preferred. In this case, a suitable dispensing device, such as the 6060 Model 6060 infusion dispenser manufactured by Baxter, can advantageously be selected. Whichever metering device is chosen, it should be of such a design or construction that minimizes or, better, eliminates the stoppage of the introduction of a therapeutic agent in the event of a replacement of the reservoir and / or replacement or recharging of the battery. This can be done, for example, by selecting a device with a second reservoir of a solution of a CD19 × CD3 bispecific single chain antibody, in addition to the reservoir that is being replaced, so that continuous infusion from this second reservoir to the patient can continue even when the empty or almost empty reservoir is removed and replaced with a new tank.

The method of intravenous administration and, as the case may be, co-administration as part of a co-therapy regimen involves the implantation of a pump into the patient's body to dose such administration. One of ordinary skill in the art will be aware of such metering pumps, such as the 6060 model manufactured by Baxter, as described above.

As a non-limiting example, it is possible that uninterrupted, i.e. continuous administration should be carried out using a small pumping system that the patient wears or which is implanted to dose the therapeutic agent into the patient. Such pumping systems are generally known in the art and are usually based on the periodic replacement of reservoirs containing a therapeutic agent administered by infusion. When replacing the reservoir in such a pumping system, a temporary interruption of the otherwise continuous flow of the therapeutic agent into the patient's body may follow. In such a case, it could still be considered, within the meaning of the pharmaceutical agents and methods of the present invention, that the administration step before replacing the reservoir and the administering step after replacing the reservoir together constitute the “continuous administration” of such a therapeutic agent. The same could be applied to very long administrations, in which the reservoir could have to be replaced more than once, or in which the batteries used to drive the pump could need to be replaced, causing the patient to temporarily stop receiving the therapeutic solution.

Appropriate measures should be taken to minimize the risk of infection at the injection site - a puncture in the patient's body, since such wounds existing over a long period of time are especially susceptible to such infection. The foregoing also applies to intramuscular administration through a similar delivery system.

Continuous administration can be transdermal by means of a patch that is on the skin and is replaced from time to time. A person skilled in the art knows plaster systems for drug delivery suitable for this purpose. It is noteworthy that transdermal administration is particularly consistent with continuous administration, since the replacement of the first depleted patch can advantageously be carried out simultaneously with the placement of a new, second patch, for example, on a skin surface area immediately adjacent to the first depleted patch and immediately before the removal of the first depleted patch. Problems associated with interruption of the flow or failure of the batteries do not occur.

In a further preferred embodiment, continuous administration is via the pulmonary route, for example, through a tube located in one or both nostrils, which is connected to a pressurized reservoir, the contents of which are accurately dosed.

In addition, the present invention relates to the construction of a CD19 × CD3 bispecific single chain antibody for treating, reducing the intensity or eliminating acute lymphoblastic leukemia (ALL) in adults. The present invention further relates to the use of the construction of a CD19 × CD3 bispecific single chain antibody for the preparation of a pharmaceutical composition for treating, reducing the intensity or eliminating acute lymphoblastic leukemia (ALL) in adults. Preferably, said acute lymphoblastic leukemia (ALL) is acute lymphoblastic leukemia from B-line cells, more preferably acute lymphoblastic leukemia from B-cell progenitors.

In a preferred embodiment of these medical applications, said acute lymphoblastic leukemia (ALL) is chemotherapy resistant in patients not suitable for allogeneic HSCT.

In an alternative embodiment of these medical applications, the administration of the CD19 × CD3 bispecific single chain antibody construct is followed by allogeneic HSCT, or these applications replace allogeneic HSCT in patients suitable for allogeneic HSCT.

In another preferred embodiment of these medical applications, the construction of a CD19 × CD3 bispecific single chain antibody is intended to treat, reduce the intensity or eliminate the minimum residual disease (MRD) in a patient with acute lymphoblastic leukemia (ALL). Preferably, when the specified patient is MRD-positive in terms of minimum residual disease with complete hematological remission.

In a further preferred embodiment of the indicated medical applications, the administration of the indicated CD19xCD3 bispecific single chain antibody will stabilize the disease or translate MRD-positive acute lymphoblastic leukemia (ALL) into an MRD-negative state.

Preferably, MRD is determined by quantifying individual rearrangements of immunoglobulin genes or rearrangements of T cell receptor (TCR) genes, or bcr / abl fusion transcripts, or t translocations (4; 11) using PCR or FACS analysis.

It is even more preferable when a patient with ALL shows a signal with respect to bcr / abl or translocation t (4; 11) that exceeds the detection limit and / or at least one rearrangement marker with sensitivity ≥10 ^-4 .

In another preferred embodiment of the indicated medical applications, the period of time until the relapse at the molecular level detected by said detection methods is more than 4 months.

In another preferred embodiment of the indicated medical applications, the corresponding variable regions of the heavy chains (V _H ) and the corresponding variable regions of the light chains (V _L ) in the indicated construction of the CD19 × CD3 bispecific single chain antibody are arranged, from the N-terminal to the C-terminal, in the following order V _L (CD19) -V _H (CD19) -V _H (CD3) -V _L (CD3).

Preferably, said CD19 × CD3 bispecific single chain antibody construct comprises an amino acid sequence reflected in SEQ ID NO: 1 or an amino acid sequence identical to SEQ ID NO: 1 by at least 90%, preferably 95%.

In a further preferred embodiment of these medical applications, one treatment cycle is continuous infusion for 4 weeks, followed by repeated cycles after a 2-week period of time without treatment.

Preferably, when the treatment cycle is repeated at least three times after determining the MRD-negative state (consolidation).

In another preferred embodiment of these medical applications, a CD19 × CD3 bispecific single chain antibody construct should be administered in a daily dose of 10 μg to 100 μg per square meter of patient surface area.

Preferably, when the design of the CD19 × CD3-bespecifically single-chain antibodies should be introduced in a daily dose of 15 μg to 30 μg per square meter of patient’s body surface area.

The definitions and explanations provided in relation to the pharmaceutical methods and means of the present invention are applicable, with necessary amendments, to medical applications of the construction of the CD19 × CD3 bispecific single chain antibody described herein.

The drawings are presented:

Figure 1. The mechanism of action of CD19 × CD3 bispecific single chain antibodies. CD19 × CD3 bispecific single chain antibody (Blinatumomab or MT103) redirects CD3 positive cytotoxic T cells to eliminate human acute lymphoblastic leukemia cells carrying the CD19 antigen.

Figure 2. An example of a course of minimal residual disease (MRD). PCR-based determination of TCR gene rearrangement (MRD) in patient 109-002 with total B-cell ALL (cALL) proves MRD positivity before treatment with CD19 × CD3 bispecific single chain antibody and persistent MRD negativity after the 1st treatment cycle of CD19 × CD3 bispecific single chain antibody.

Figure 3. The dynamics of CD4 and CD8 T cells of the patient 109-002 during cycle 1 treatment. A representative example of pharmacodynamics, demonstrating the rapid redistribution of T cells and an increase, mainly, the number of cytotoxic CD8 T cells.

Figure 4. The dynamics of the T-cell subpopulations of patient 109-002 during cycle 1 of treatment. A representative example of pharmacodynamics, demonstrating the rapid redistribution of T cells and the multiplication of memory cells - T-effectors (TEM). Untrained T cells do not multiply.

Figure 5. The first four patients included in the phase II study. All patients had previously been prescribed standard chemotherapy regimens for ALL according to GMALL protocols, including at least one treatment for consolidation.

6. Minimum residual disease (MRD) responses in these patients with ALL (i.e., the first four patients included in the phase II study) after the first cycle of treatment with a CD19 × CD3 bispecific single chain antibody.

7. Currently adjusted minimum residual disease (MRD) response data. MRD negativity could be achieved in nine of eleven patients with rearrangements of immunoglobulin or TCR genes, in one of two patients with t translocations (4; 11), and in three of four patients with bcr / abl transcripts. As a result, 13 of 16 measurable patients (81%) became MRD-negative.

Fig. 8. The period of MRD-negativity (state as of May 25, 2009). The longest period of MRD negativity observed so far in patient 108-001, not transplanted after treatment with antibody, is 41 weeks. Patient 111-001, who was MRD-negative from 06/23/2008 to 10/27/2008 after treatment with a CD19 × CD3 bispecific single chain antibody and who successfully transplanted allogeneic hematopoietic stem cells thereafter, has not suffered a relapse so far. An arrow indicates that the response is still persisting (as of May 25, 2009). Patient 109-002 (*) suffered a testicular relapse followed by hematologic relapse after 19 weeks of MRD negativity.

The present invention is further illustrated by the following example.

Example

1. The creation, expression and cytotoxic activity of a CD19 × CD3 bispecific single chain antibody are described in WO 99/54440. The corresponding amino acid sequence and nucleic acid sequence of a CD19 × CD3 bispecific single chain antibody are indicated in SEQ ID NOs: 1 and 2, respectively. VH and VL regions of the CD3 binding domain of a CD19 × CD3 bispecific single chain antibody are indicated in SEQ ID NOs: 7-10, respectively, while VH and VL regions of the CD19 binding domain of a CD19 × CD3 bispecific single chain antibody are shown in SEQ ID NO: 3-6, respectively. The corresponding CDR plots are presented in SEQ ID NO: 11-22.

2. The ongoing phase I test in patients with recurrent B-cell non-Hodgkin lymphoma (NHL) shows a high degree of response at a dose of 60 μg / m ² / day of CD19 × CD3 bispecific single chain antibody. The duration of responses is up to more than 12 months (persisting in several patients). Removal of bone marrow-infiltrating B-NHL cells began at a dose of 15 μg / m ² / day (Bargou et al., Science 2008).

3. Based on these results, a phase II dose increase study was planned in conjunction with the German Multicenter Study Group on Adult Acute Lymphoblastic Leukemia (GMALL) to study the efficacy, safety and tolerability of CD19 × CD3 bispecific single chain antibodies in adult patients with acute lymphoblastic leukemia (ALL ) (not transplanted) that achieved complete hematologic remission but still remained MRD-positive. MRD is an independent prognostic factor that reflects resistance to the original drug (s) and is associated with a high risk of relapse after the onset of consolidation. This applies to Ph + / BCR-ABL-positive and -negative ALL. MRD was determined using standard methods, either by quantifying individual rearrangements of immunoglobulin genes or T cell receptors (TCR), or by detecting bcr / abl fusion transcripts or t translocations (4; 11). The study population includes adult patients with acute lymphoblastic leukemia (ALL) from precursor B cells that show a signal for bcr / abl or t (4; 11) that exceeds the detection limit and / or at least one marker for rearrangement with sensitivity ≥10 ^-4 . More specifically, the main inclusion criteria included:

- Patients with ALL from precursor B cells in complete hematological remission with impaired or relapse at the molecular level starting at any time after consolidation 1 from first-line therapy in standard protocols.

- Patients should have a molecular marker to assess the minimum residual disease, which is either bcr / abl or translocation t (4; 11) at any detection level, or separate rearrangements of immunoglobulin or TCR genes determined by analysis with a sensitivity of at least 10 ^-4 and quantitative limits up to 10 ^-4 for at least one marker.

The initial expected result of a (conducting) Phase II study is the degree of transition to a negative state according to the indicator of minimum residual disease (MRD), determined by a signal with respect to bcr / abl or translocation t (4; 11), which is below the detection limit, and / or the identification of individual gene rearrangements of immunoglobulins or T-cell receptors (TCR) at a level that is below 10 ^-4 . Secondary expected results are the period of time before hematologic relapse, the period of time before the progression of MRD, and the period of time before relapse at the molecular level. One treatment cycle for a CD19 × CD3 bispecific single chain antibody is a 4-week continuous intravenous infusion, followed by transplantation of allogeneic hematopoietic stem cells after the first cycle or further cycles, or repeated cycles after a 2-week period without treatment. The state of minimum residual disease (MRD) is monitored after each treatment cycle. The initial dose level of 15 microgram / m ^2/24 hours, but it may be increased up to 30 micrograms / m ^2/24 h, and higher dose levels (60 micrograms / m ^2/24 hours or 90 micrograms / m ^2/24 hours ) based on clinical activity and safety data. For statistical calculation, MinMax uses a two-stage calculation of Simon (14 out of 21 patients).

Further, the data on the first four patients included in the study are presented in more detail as an example. These four patients age 31, 57, 62 and 65 was assigned an initial dose level is 15 mcg / m ^2/24 hours. As shown in Figure 5, patients with a number 111-001, 109-002 and 110-002 general B-cell ALL (cALL) was diagnosed, while patient No. 108-001 is a patient with pre-B-cell ALL. Four patients were previously prescribed standard chemotherapy regimens for ALL according to the GMALL protocols, including at least one treatment for consolidation. All of them were resistant to chemotherapy with regard to minimum residual disease (MRD). More specifically, all patients were MRD-positive with complete hematologic remission. Patients Nos. 110-002, 108-001, and 109-002 were not suitable for transplantation of allogeneic hematopoietic stem cells, while patients with No. 111-0001 were suitable for this transplantation.

As shown in Fig.6, three of the first 4 patients included in the study had a minimal residual disease (MRD), determined by rearrangements of immunoglobulin or TCR genes at levels of 10 ^-4 (in the case of a patient with No. 111-001), 10 ^-3 (in the case of the patient with No. 108-001) and 10 ^-1 (in the case of the patient with No. 109-002), and one patient (patient with No. 110-002) had an MRD determined by the bcr / abl fusion transcripts on level of 10 ^-4 . Three out of 3 patients, i.e. patients No. 111-001, 108-001, and 109-002, with rearrangements of immunoglobulin or TCR genes, were MRD negative after the first treatment cycle, regardless of the level of MRD positivity in the baseline. Patient No. 111-001, the only one of four patients suitable for transplantation of allogeneic hematopoietic stem cells, underwent transplantation after being converted to an MRD negative state as a result of treatment with a CD19 × CD3 bispecific single chain antibody.

Figure 2 presents an example of the course of the minimum residual disease (MRD) in a patient 109-002. PCR-based determination of TCR gene rearrangement (MRD) in patient 109-002 with common B-cell ALL proves MRD positivity prior to treatment with CD19 × CD3 bispecific single chain antibody (blinatumomab) and MRD negativity starting after the 1st treatment cycle of CD19 × CD3 bispecific single chain antibody and lasting up to week 19. After this, the patient underwent testicular relapse followed by hematological relapse.

Another patient, numbered 110002, had a steady bcr / abl level with no signs of hematologic relapse after the initial treatment cycle; see Fig.6.

The treatment of patients with CD19 × CD3 bispecific single chain antibody was well tolerated: with the exception of fever in the first 3 days of treatment, clinically significant toxicities were not recorded.

Meanwhile, for this number seventeen adult patients have been treated, or are still being treated, with a CD19 × CD3 bispecific single chain antibody. All patients were resistant to conventional ALL therapy, including chemotherapy, prior to antibody treatment. Prior to antibody treatment, none of them allogeneic hematopoietic stem cells were transplanted. The average age of the patients was 48 years, ranging from 20 to 77 years. Ten patients were women, seven patients were men. 14 patients were assigned dose level is 15 mcg / m ^2/24 hours CD19 × CD3-bispecific single chain antibody, whereas three patients the dose was increased from 15 to 30 microgram / m ^2/24 hours after the first or further treatment cycles: in the case of patient 109-004, the dose was increased after the second treatment cycle (using only three treatment cycles followed by transplantation of allogeneic hematopoietic stem cells), in the case of patient 109-003, after the third treatment cycle (using only four treatment cycles), and in the case of patient 110-002, after the sixth treatment cycle (using a total of seven treatment cycles). Eleven of these patients had minimal residual disease (MRD) as determined by rearrangements of immunoglobulin or TCR genes, two patients had t translocations (4; 11), and four patients showed bcr / abl fusion transcripts.

As a result, the MRD response was measurable in 16 of 17 patients. As shown in FIGS. 7, 13 of 16 measurable patients became MRD-negative, corresponding to an amazing degree of complete response at the molecular level of 81%. More specifically, MRD negativity could be achieved in nine of eleven patients with rearrangements of immunoglobulin or TCR genes, in one of two patients with t translocations (4; 11), and in three of four patients with bcr / abl transcripts. As shown in FIG. 8, the longest period of MRD negativity observed so far in patient 108-001 not transplanted after treatment with antibody is 41 weeks. Another patient, who was MRD-negative from 06/23/2008 to 10/27/2008 and who successfully transplanted allogeneic hematopoietic stem cells after treatment with an antibody, has not suffered a relapse so far, see patient 111-001 in Fig. 8. It is noteworthy that patients with bcr / abl, who could be successfully treated with a CD19 × CD3 bispecific single chain antibody, in the previous ALL treatment regimen were resistant to imatinib and / or dasatinib tyrosine kinase inhibitors and not tolerating them. In particular, one of the bcr / abl patients who responded to treatment with a CD19 × CD3 bispecific single chain antibody had a T315I mutation that was resistant to therapy with tyrosine kinase inhibitors. Thus, the introduction of a CD19 × CD3 bispecific single chain antibody is now the first to provide therapy for dasatinib-resistant ALL patients with bcr / abl transcripts. Only three of the total 17 patients did not become MRD-negative. However, in two of them, stabilization of the disease could be achieved. Only one patient with an initial stable disease suffered a hematologic relapse in the third cycle of treatment. One patient could not be evaluated due to a serious side effect on day 2 of the study.

In conclusion, a definitely exceptional degree of complete response at the molecular level of 81% could be achieved in patients with ALL from precursor B cells as a result of treatment with a CD19 × CD3 bispecific single chain antibody. The activity of this antibody could be registered in all subgroups of patients treated, including in patients resistant to tyrosine kinase inhibitors (T315I) with bcr / abl and patients with translocations t (4; 11). In addition, when treating a CD19 × CD3 bispecific single chain antibody, a suitable toxicity profile is demonstrated, as opposed to conventional ALL therapies such as chemotherapy. In view of this, the administration of the CD19 × CD3 bispecific single chain antibody described herein provides a new and advantageous treatment option for acute lymphoblastic leukemia (ALL), especially in cases where ALL is resistant to conventional therapy for ALL, such as chemotherapy. In addition, the introduction of a CD19 × CD3 bispecific single chain antibody is now the first to determine therapy for MRD-positive ALL.

These currently corrected results show that treatment of patients with acute lymphoblastic leukemia (ALL) with CD19 × CD3 bispecific single chain antibody can translate acute minimal lymphoblastic leukemia (ALL) positive in terms of minimum residual disease (MRD) into MRD-negative state (as shown as an example, in ALL patients suffering from immunoglobulin or TCR gene rearrangements, bcr / abl transcripts, or t translocations (4; 11), and that this treatment is well tolerated. In view of this, the introduction of CD19 × CD3 bispecific The single chain antibody described herein provides an alternative treatment option, in particular acute lymphoblastic leukemia (ALL) in adults, especially ALL, resistant to conventional therapy for ALL, such as chemotherapy and / or hematopoietic stem cell transplantation. Treatment of CD19 × CD3 A b-specific single chain antibody is particularly preferred for the treatment of MRD-positive ALL.

Claims (21)

1. The use of blinatumomab (MT103) to obtain a pharmaceutical composition for treating, reducing the intensity or eliminating acute lymphoblastic leukemia (ALL), where blinatumomab (MT103) translates MRD (minimal residual disease) - positive acute lymphoblastic leukemia ALL to MRD-negative state ALL. 2. The use according to claim 1, wherein said acute lymphoblastic leukemia (ALL) is acute lymphoblastic leukemia (ALL) from B-cell cells. 3. The use according to claim 2, wherein said acute lymphoblastic leukemia from B-line cells is acute lymphoblastic leukemia from B-precursor cells. 4. The use according to any one of paragraphs. 1-3, in which the specified acute lymphoblastic leukemia (ALL) is resistant to chemotherapy in patients not suitable for transplantation of allogeneic hematopoietic stem cells. 5. The use according to any one of paragraphs. 1-3, followed by transplantation of allogeneic hematopoietic stem cells or which replaces transplantation of allogeneic hematopoietic stem cells in patients suitable for transplantation of allogeneic hematopoietic stem cells. 6. The use according to any one of paragraphs. 1-3, which is intended to treat, reduce the intensity or eliminate the minimum residual disease (MRD) in a patient with acute lymphoblastic leukemia (ALL). 7. The use according to claim 6, in which the specified patient is MRD-positive with complete hematological remission. 8. The use according to claim 7, in which the MRD is determined by quantifying individual rearrangements of immunoglobulin genes or rearrangements of T cell receptor (TCR) genes, or bcr / abl fusion transcripts, or t translocations (4; 11) using PCR analysis or FACS. 9. The use of claim 8, in which the patient with ALL shows a signal for bcr / abl or translocation t (4; 11) that exceeds the detection limit, and / or at least one rearrangement marker with sensitivity ≥10 ^-4 . 10. The use according to any one of paragraphs. 1-3, 7, 9, in which the period of time before relapse at the molecular level, determined by quantifying individual rearrangements of immunoglobulin genes or rearrangements of T-cell receptor (TCR) genes, or bcr / abl fusion transcripts, or t translocations (4 ; 11) using PCR or FACS analysis, is more than 4 months. 11. The use according to any one of paragraphs. 1-3, 7, 9, in which the corresponding variable regions of the heavy chains (V _H ) and the corresponding variable regions of the light chains (V _L ) in the indicated pancreaticomab (MT 103) are located, from the N-end to the C-end, in the following order V _L (CD19) -V _H (CD19) -V _H (CD3) -V _L (CD3). 12. The use according to claim 11, in which the specified blinatumomab (MT 103) includes the amino acid sequence reflected in SEQ ID NO: 1. 13. The use according to any one of paragraphs. 1-3, 7, 9, in which one treatment cycle is a continuous administration for 4 weeks. 14. The application of item 13, in which the specified treatment cycle is followed by repeated cycles after a 2-week period of time without treatment. 15. The use of claim 14, wherein said treatment cycle must be repeated at least two, three, four, five, six, seven, eight, nine or ten times. 16. The use according to any one of paragraphs. 14-15, in which the treatment cycle is repeated at least three times after determining the MRD-negative state. 17. The use according to any one of paragraphs. 1-3, 7, 9, 12, 14, 15, in which blinatumomab (MT103) should be administered in a daily dose of 10 μg to 100 μg per square meter of patient’s body surface area. 18. The application of clause 17, in which blinatumomab (MT103) should be administered in a daily dose of 15 μg to 30 μg per square meter of patient’s body surface area. 19. The use according to claim 17, wherein the pancreaticomab (MT103) should be administered in an amount of 5 μg on the first day (s) of the infusion period, followed by the administration of 15 μg per square meter of the patient’s body surface for the remaining treatment period. 20. The use according to claim 17, wherein the pancreaticomab (MT103) should be administered in an amount of 15 μg on the first day (s) of the infusion period, followed by 30 μg or 45 per square meter of the patient’s body surface during the remaining treatment period. 21. The use according to claim 17, wherein the pancreaticomab (MT103) is to be administered in an amount of 5 μg on the first day (s) of the infusion period, followed by the administration of 15 μg on the subsequent day (s) of the infusion period, followed by the introduction of 45 μg per square meter of surface area of the patient’s body as a daily dose for the remaining treatment period.

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